Prof Marina Drepano

Prévia do material em texto

T h e n e w e ngl a nd j o u r na l o f m e dic i n e
n engl j med 376;9 nejm.org March 2, 2017848
Brief Report
Summ a r y
Sickle cell disease results from a homozygous missense mutation in the β-globin 
gene that causes polymerization of hemoglobin S. Gene therapy for patients with 
this disorder is complicated by the complex cellular abnormalities and challenges 
in achieving effective, persistent inhibition of polymerization of hemoglobin S. We 
describe our first patient treated with lentiviral vector–mediated addition of an 
antisickling β-globin gene into autologous hematopoietic stem cells. Adverse events 
were consistent with busulfan conditioning. Fifteen months after treatment, the 
level of therapeutic antisickling β-globin remained high (approximately 50% of 
β-like–globin chains) without recurrence of sickle crises and with correction of the 
biologic hallmarks of the disease. (Funded by Bluebird Bio and others; HGB-205 
ClinicalTrials.gov number, NCT02151526.)
Sickle cell disease is among the most prevalent inherited mono-genic disorders. Approximately 90,000 people in the United States have sickle cell disease, and worldwide more than 275,000 infants are born with 
the disease annually.1,2 Sickle cell disease was the first disease for which the mo-
lecular basis was identified: a single amino acid substitution in “adult” βA-globin 
(Glu6Val) stemming from a single base substitution (A→T) in the first exon of the 
human βA-globin gene (HBB) was discovered in 1956.3 Sickle hemoglobin (HbS) 
polymerizes on deoxygenation, reducing the deformability of red cells. Patients 
have intensely painful vaso-occlusive crises, leading to irreversible organ damage, 
poor quality of life, and reduced life expectancy. Hydroxyurea, a cytotoxic agent 
that is capable of boosting fetal hemoglobin levels in some patients, is the only dis-
ease-modifying therapy approved for sickle cell disease.4
Allogeneic hematopoietic stem-cell transplantation currently offers the only cura-
tive option for patients with severe sickle cell disease.5,6 However, fewer than 18% 
of patients have access to a matched sibling donor.7,8 Therapeutic ex vivo gene trans-
fer into autologous hematopoietic stem cells, referred to here as gene therapy, may 
provide a long-term and potentially curative treatment for sickle cell disease.9
We previously reported proof of effective, sustained gene therapy in mouse mod-
The authors’ affiliations are listed in the 
Appendix. Address reprint requests to 
Dr. Cavazzana at the Biotherapy Depart-
ment, Necker Children’s Hospital, Assis-
tance Publique–Hôpitaux de Paris, 149 
rue de Sèvres, 75015 Paris, France, or at 
 m . cavazzana@ aphp . fr; or to Dr. Leboulch 
at the Institute of Emerging Diseases and 
Innovative Therapies, 18, rte du Panorama 
BP-6, 92265 Fontenay-aux-Roses, France, 
or at pleboulch@ rics . bwh . harvard . edu.
Drs. Ribeil and Hacein-Bey-Abina and 
Drs. Leboulch and Cavazzana contributed 
equally to this article
N Engl J Med 2017;376:848-55.
DOI: 10.1056/NEJMoa1609677
Copyright © 2017 Massachusetts Medical Society.
Gene Therapy in a Patient 
with Sickle Cell Disease
Jean-Antoine Ribeil, M.D., Ph.D., Salima Hacein-Bey-Abina, Pharm.D., Ph.D., 
Emmanuel Payen, Ph.D., Alessandra Magnani, M.D., Ph.D., 
Michaela Semeraro, M.D., Ph.D., Elisa Magrin, Ph.D., Laure Caccavelli, Ph.D., 
Benedicte Neven, M.D., Ph.D., Philippe Bourget, Pharm.D., Ph.D., 
Wassim El Nemer, Ph.D., Pablo Bartolucci, M.D., Ph.D., Leslie Weber, M.Sc., 
Hervé Puy, M.D., Ph.D., Jean-François Meritet, Ph.D., David Grevent, M.D., 
Yves Beuzard, M.D., Stany Chrétien, Ph.D., Thibaud Lefebvre, M.D., 
Robert W. Ross, M.D., Olivier Negre, Ph.D., Gabor Veres, Ph.D., 
Laura Sandler, M.P.H., Sandeep Soni, M.D., Mariane de Montalembert, M.D., Ph.D., 
Stéphane Blanche, M.D., Philippe Leboulch, M.D., and Marina Cavazzana, M.D., Ph.D. 
The New England Journal of Medicine 
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n engl j med 376;9 nejm.org March 2, 2017 849
Brief Report
els of sickle cell disease by lentiviral transfer of 
a modified HBB encoding an antisickling variant 
(βA87Thr:Gln [βA-T87Q]).10,11 Here we report the re-
sults for a patient who received lentiviral gene 
therapy in the HGB-205 clinical study and who 
had complete clinical remission with correction of 
hemolysis and biologic hallmarks of the disease.
C a se R eport
A boy with the βS/βS genotype, a single 3.7-kb 
α-globin gene deletion, and no glucose 6-phos-
phate dehydrogenase deficiency received a diag-
nosis of sickle cell disease at birth and was fol-
lowed at the Reference Centre for Sickle Cell 
Disease of Necker Children’s Hospital in Paris. He 
had a history of numerous vaso-occlusive crises, 
two episodes of the acute chest syndrome, and bi-
lateral hip osteonecrosis. He had undergone cho-
lecystectomy and splenectomy. During screening, 
a cerebral hypodensity without characteristics of 
cerebral vasculopathy was detected.
Because hydroxyurea therapy administered 
when the boy was between 2 and 9 years of age did 
not reduce his symptoms significantly, a prophy-
lactic red-cell transfusion program was initiated 
in 2010, including iron chelation with deferasirox 
(at a dose of 17 mg per kilogram of body weight 
per day). He had had an average of 1.6 sickle cell 
disease–related events annually in the 9 years be-
fore transfusions were initiated.
In May 2014, he was enrolled in our clinical 
study. His verbal assent and his mother’s written 
informed consent were obtained. In October 2014, 
when the patient was 13 years of age, he received 
an infusion of the drug product LentiGlobin 
BB305.
Me thods
Study Oversight
The study protocol, which is available with the 
full text of this article at NEJM.org, was designed 
by the last two authors and Bluebird Bio, the 
study sponsor. The protocol was reviewed by the 
French Comité de Protection des Personnes and 
relevant institutional ethics committees. Clinical 
data were collected by the first author, and labo-
ratory data were generated by the sponsor, the last 
author, and other authors. The authors had access 
to all data, and data analysis was performed by 
them. The first author and one author employed 
by the sponsor wrote the first draft of the man-
uscript, which was substantively revised by the 
last two authors and further edited and approved 
by all the authors with writing assistance provided 
by an employee of the sponsor. The authors vouch 
for the accuracy and completeness of the data and 
adherence to the protocol.
Antisickling Gene Therapy Vector
The structure of the LentiGlobin BB305 vector has 
been previously described (see Fig. S1 in the Sup-
plementary Appendix, available at NEJM.org).12 
This self-inactivating lentiviral vector encodes the 
human HBB variant βA-T87Q. In addition to inhibit-
ing HbS polymerization, the T87Q substitution 
allows for the β-globin chain of adult hemoglo-
bin (HbA)T87Q to be differentially quantified by 
means of reverse-phase high-performance liquid 
chromatography.12
Gene Transfer and Transplantation 
Procedures
Bone marrow was obtained twice from the pa-
tient to collect sufficient stem cells for gene trans-
fer and backup (6.2×108 per kilogram and 5.4×108 
per kilogram, respectively, of total nucleated cells 
obtained). Both procedures were preceded by ex-
change transfusion, and bone marrow was ob-
tained without clinical sequelae. Anemia was the 
only grade 3 adverse event reported during these 
procedures. Bone marrow–enriched CD34+ cells 
were transduced with LentiGlobin BB305 vector 
(see the Methods section in the Supplementary 
Appendix).13 The mean vector copy numbers for 
the two batches of transduced cells were 1.0 and 
1.2 copies per cell.
The patient underwent myeloablation with in-
travenous busulfan (see the Methods section in the 
SupplementaryAppendix). The total busulfan area 
under the curve achieved was 19,363 μmol per 
minute. After a 2-day washout period, transduced 
CD34+ cells (5.6×106 CD34+ cells per kilogram) 
were infused. Red-cell transfusions were to be 
continued after transplantation until a large pro-
portion of HbAT87Q (25 to 30% of total hemoglo-
bin) was detected.
The patient was followed for engraftment; toxic 
effects (graded according to the National Cancer 
Institute Common Terminology Criteria for Ad-
verse Events, version 4.03); vector copy number in 
total nucleated blood cells and in different lin-
eages; quantification of HbAT87Q, HbS, and fetal 
hemoglobin levels by means of high-performance 
liquid chromatography; DNA integration-site map-
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n engl j med 376;9 nejm.org March 2, 2017850
T h e n e w e ngl a nd j o u r na l o f m e dic i n e
ping by linear amplification–mediated polymerase 
chain reaction in nucleated blood cells; and repli-
cation-competent lentivirus analysis by p24 anti-
body enzyme-linked immunosorbent assay. Red-
cell analyses were performed at month 12 (see the 
Methods section in the Supplementary Appendix).
R esult s
Engraftment and Gene Expression
Neutrophil engraftment was achieved on day 38 
after transplantation, and platelet engraftment was 
achieved on day 91 after transplantation. Figure 1A 
shows the trajectory of vector copy numbers and 
Figure 1B shows production of HbAT87Q. Gene 
marking increased progressively in whole blood, 
CD15 cells, B cells, and monocytes (Fig. S2 in the 
Supplementary Appendix), stabilizing 3 months 
after transplantation. Increases in levels of vector-
bearing T cells were more gradual.
HbAT87Q levels also increased steadily (Fig. 1B) 
and red-cell transfusions were discontinued, with 
the last transfusion on day 88. Levels of HbAT87Q 
reached 5.5 g per deciliter (46%) at month 9 and 
continued to increase to 5.7 g per deciliter (48%) 
at month 15, with a reciprocal decrease in HbS 
levels to 5.5 g per deciliter (46%) at month 9 and 
5.8 g per deciliter (49%) at month 15. Total he-
moglobin levels were stable between 10.6 and 
12.0 g per deciliter after post-transplantation 
Figure 1. Engraftment with Transduced Cells and Therapeutic Gene Expression in the Patient.
Panel A shows vector copy number values in blood nucleated cells and the short-lived CD15+ (neutrophils) fraction 
thereof over 15 months after infusion of transduced CD34+ cells. Initial values in transduced cells before the infu-
sion are shown. Panel B shows total hemoglobin levels and calculated levels of each hemoglobin fraction based on 
high-performance liquid chromatography measurements of globin chains. The percent contribution of hemoglobin 
fractions at month 15 is also indicated. The hemoglobin A (HbA) levels are derived from the regular red-cell trans-
fusions received by the patient before gene therapy and briefly thereafter (the last red-cell transfusion occurred on 
day 88). HbA2 is an alternative adult hemoglobin that is not derived from transfused blood. HbF denotes fetal he-
moglobin, and HbS sickle hemoglobin.
V
ec
to
r 
C
op
y 
N
o.
/D
ip
lo
id
 G
en
om
e 3.0
2.0
2.5
1.5
1.0
0.5
0.0
0 3 6 9 12 15
Months after Infusion of Transduced CD34+ Cells
B
A
Vector copy no. in
transduced CD34+ cells
before infusion
Peripheral blood
CD15+ cells
H
em
og
lo
bi
n 
C
on
ce
nt
ra
tio
n
(g
/d
l)
15.0
10.0
5.0
0.0
0 3 6 9 12 15
Months after Infusion of Transduced CD34+ Cells
HbAT87Q 48% 
HbS 49% HbA
Total hemoglobin 11.8 g/dl
HbA2 2% 
HbF <1% 
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n engl j med 376;9 nejm.org March 2, 2017 851
Brief Report
month 6. Fetal hemoglobin levels remained below 
1.0 g per deciliter.
Safety
The patient had expected side effects from busulfan 
conditioning. Grade 3 and 4 events included grade 
4 neutropenia, grade 3 anemia, grade 3 thrombo-
cytopenia, and grade 3 infection with Staphylococcus 
epidermidis (with positive results on blood culture), 
all of which resolved with standard measures. Af-
ter the patient was discharged from the hospital, 
four grade 2 adverse events were reported: lower 
limb pain 3 months after treatment and transient 
increases in alanine aminotransferase, aspartate 
aminotransferase, and γ-glutamyltransferase be-
tween 5 and 8 months after treatment. All these 
events resolved spontaneously.
No adverse events related to the LentiGlobin 
BB305–transduced stem cells were reported (Ta-
ble S1 in the Supplementary Appendix). Test results 
for the presence of replication-competent lentivi-
rus were uniformly negative. Serial monitoring of 
integration sites in peripheral-blood samples 
showed a consistently polyclonal profile without 
detection of a dominant clone (defined as a single 
clone accounting for >30% of unique integration 
events) through month 12 (Fig. S3 in the Supple-
mentary Appendix).
Clinical and Biologic Measures
The patient was discharged on day 50. More than 
15 months after transplantation, no sickle cell 
disease–related clinical events or hospitalization 
had occurred; this contrasts favorably with the 
period before the patient began to receive regu-
lar transfusions. All medications were discontin-
ued, including pain medication. The patient re-
ported full participation in normal academic and 
physical activities. Magnetic resonance imaging 
(MRI) of the head at 8 months showed unchanged 
punctate subcortical white-matter hypodensities. 
Lower limb MRI at 14 months showed no recent 
bone or tissue damage.
Changes in sickle cell disease–related biologic 
measures are shown in Table 1. Complete blood 
counts were stable, reticulocyte counts decreased 
substantially (Fig. S4 in the Supplementary Appen-
dix), and circulating erythroblasts were not de-
tected. Laboratory values, including urinary micro-
albumin levels, indicated normal renal and liver 
functions. Although iron chelation was discontin-
ued before transplantation, the ferritin levels de- T
ab
le
 1
. K
ey
 L
ab
or
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26
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9
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 (
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 (
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 Ex
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 w
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 d
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 n
ot
 d
on
e.
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n engl j med 376;9 nejm.org March 2, 2017852
T h e n e w e ngl a nd j o u r na l o f m e dic i n e
Si
ck
le
d 
R
ed
 C
el
ls
 (%
)
15 60
5
10
0
Pa
tie
nt
6 M
o
Pa
tie
nt
12
 M
o
Co
nt
ro
l 1
Co
nt
ro
l 2
Co
nt
ro
l 3
Co
nt
ro
l 4
Co
nt
ro
l 5
C Oxygen Equilibrium Curves (37°C, pH 7.4)
A Sickled Red Cells, Normoxic Conditions
Si
ck
le
d 
R
ed
 C
el
ls
 (%
)
20
40
0
Pa
tie
nt
6 M
o
Pa
tie
nt
12
 M
o
Co
nt
ro
l 1
Co
nt
ro
l 2
Co
nt
ro
l 3
Co
nt
ro
l 4
Co
nt
ro
l 5
B Sickled Red Cells, Hypoxic Conditions
O
xy
ge
n 
Fr
ac
tio
na
l S
at
ur
at
io
n 
(%
) 1.0
0.6
0.4
0.2
0.8
0.0
0 20 40 60 80 100 120 140
Partial Pressure of Oxygen (mm Hg)
Phthalate Density (mg/ml)
D Red-Cell Deformability
E Red-Cell Density
El
on
ga
tio
n 
In
de
x
0.7
0.5
0.4
0.2
0.3
0.1
0.6
0.0
R
ed
 C
el
ls
 (%
)
100
60
40
20
80
90
50
30
10
70
0
1.075 1.080 1.085 1.090 1.095 1.100 1.105 1.110 1.115 1.120
0.30 0.53 0.95 1.69 3.00 5.33 9.49 16.87 30.00
Shear Force (Pa) 
Patient, deoxygenation
Patient, reoxygenation
Control 1, deoxygenation
Patients with sickle cell disease,
deoxygenation
Patients with sickle cell disease,
reoxygenation
1 2
3
4
5
6
7
8
9 10
Patient 12 mo after gene therapy
Control 1
Control 6
Mean curve for healthy
participants
Low
Medium
High
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n engl j med 376;9 nejm.org March 2, 2017 853
Brief Report
creased to 363 μg per liter at month 15, and MRI 
of the liver 1 year after treatment showed a low 
iron load (level of mobilizable circulating iron, 
relaxation rate R2* = 117 Hz; and iron level, 3.1 mg 
per gram vs. 54 Hz and 14.6 mg per gram before 
gene therapy). Plasma levels of total bilirubin and 
lactate dehydrogenase normalized. Soluble trans-
ferrin receptor levels improved and were 3.4 times 
as high as normal values at screening and 1.5 
times as high at months 12 and 15, indicating 
progressive normalization of erythropoiesis.
Because the patient received a regular trans-
fusion regimen for 4 years before this study and 
because of the exchange transfusion before trans-
plantation, meaningful comparative studies before 
and after transplantation could not be conducted. 
However, the proportions of sickled red cells in 
the patient’s blood at months 6 and 12 were sig-
nificantly lower than those in untreated patients 
with sickle cell disease (βS/βS) (Fig. 2A). At 
month 12, the sickling rate in hypoxic conditions 
was not significantly different from that of the 
patient’s asymptomatic, heterozygous (βA/βS) moth-
er (Fig. 2B). Oxygen dissociation studies, which 
quantify oxygen saturation relative to the partial 
pressure of oxygen, showed that results in the 
patient at month 12 and results in a heterozygous 
(βA/βS) control were similar (Fig. 2C and 2D).
Discussion
This case report of a patient with sickle cell disease 
who received gene therapy with the use of lentiviral 
gene addition of an antisickling β-globin variant 
provides proof of concept for this approach and 
may help to guide the design of future clinical 
trials of gene therapy for sickle cell disease. Once 
the transduced stem cells engrafted, normal blood-
cell counts were ultimately attained in all lineages. 
Increasing levels of vector-bearing nucleated cells 
in the blood over the first 3 months after trans-
plantation and general vector copy number sta-
bility through month 15 suggest engraftment of 
transduced stem cells that were capable of long-
term repopulation. No adverse events that were 
considered by the investigators to be related to the 
BB305-transduced cells were observed, and the 
pattern of vector integration remained polyclonal 
without clonal dominance.14
Insertional oncogenesis has been reported in 
clinical gene-transfer studies with gamma retro-
viral vectors but not with lentiviral vectors.15,16 Un-
like gamma retroviruses, lentivirus tends to insert 
in transcriptionally active regions rather than near 
transcriptional start sites.17 In addition, the BB305 
vector is an enhancer-deleted vector and is self-
inactivating.12 Reported data from this and other 
ongoing studies of the BB305 vector involving 
patients with sickle cell disease (7 patients) and 
β-thalassemia (22 patients) show a consistent 
safety profile, with no evidence of insertional mu-
tagenesis through 4 to 30 months.18,19
Figure 2 (facing page). Results of Sickle Cell Disease–
Specific Red-Cell Assays.
Panel A shows rates of red-cell sickling under normoxic 
conditions (20% oxygen saturation) and Panel B shows 
rates of red-cell sickling under hypoxic conditions 
(10% oxygen saturation) in the patient at 6 months 
and 12 months after gene therapy and among control 
patients from whom red-cell samples were obtained: 
two patients with heterozygous A/S “sickle trait” 
(Controls 1 and 2; Control 1 is the patient’s mother) 
and three patients with sickle cell disease (Controls 3, 
4, and 5). Similar results were obtained at 7% and 5% 
oxygen saturation rates (data not shown). T bars indi-
cate standard errors. Panel C shows oxygen dissocia-
tion curves for red cells 12 months after gene therapy 
in the patient and in the patient’s heterozygous (A/S) 
mother (Control 1). These analyses were performed 
 simultaneously, under identical conditions. The mean 
red-cell deoxygenation curve (solid black line) and the 
mean red-cell reoxygenation curve (dashed black line) 
for 15 untreated patients with sickle cell disease are 
also shown. Panel D shows red-cell deformability 12 
months after gene therapy in the patient as compared 
with his heterozygous (A/S) mother (Control 1) and 
another patient with sickle cell disease (Control 6). 
The gray zone demarcates the range within which 95% 
of non–sickle cell disease red cells fall, and the black 
curve is the mean curve for healthy participants. The 
elongation index was calculated as the ratio of the 
length (A) and width (B) of a cell, where (A−B) was 
 divided by (A+B), and the result was expressed as a 
decimal between 0 and 1. Panel E shows the red-cell 
density profile 12 months after gene therapy in the 
 patient, obtained with the use of a phthalate gradient. 
We measured 10 samples (indicated with the numbers 
1 through 10 on the black curve) at various phthalate 
densities. Red lines demarcate three different densi-
ties of cells: low (<1.086 mg per milliliter), medium 
(1.086 to 1.096 mg per milliliter), and high (>1.096 mg 
per milliliter). Orange lines indicate limits of a normal 
profile. The values for the patient are shifted to the 
left because of the associated single α-globin gene 
 deletion. Cells denser than 1.110 mg permilliliter of 
phthalate solution are considered to be dense cells.
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n engl j med 376;9 nejm.org March 2, 2017854
T h e n e w e ngl a nd j o u r na l o f m e dic i n e
The appearance of vector-bearing cells in the 
periphery corresponds to the time frame for 
engraftment of long-term progenitors and stem 
cells repopulating the space of nucleated cells. 
In contrast, the slower pace for the increase of 
HbAT87Q expression reflects the more gradual time 
course of replacement of transfused red cells from 
the pretransplantation and peritransplantation pe-
riods by newly matured, graft-derived red cells.
In mouse models of sickle cell disease, thera-
peutic globin expression after gene addition was 
difficult to obtain, presumably because of competi-
tion with endogenous β-globin messenger RNAs.11 
In the current study, a high concentration of thera-
peutic HbAT87Q (ratio of HbAT87Q to HbS, approxi-
mately 1) was achieved.10,11 HbAT87Q expression 
appears to be sufficient to suppress hemolysis, 
resulting in stable hemoglobin concentrations of 
11 to 12 g per deciliter and major improvement 
in all measurable sickle cell disease–specific bio-
logic markers and blocking sickle cell disease–
related clinical events.20,21
Additional data on LentiGlobin treatment in 
sickle cell disease is currently being collected in 
HGB-206, a multicenter, phase 1/2 clinical study 
in the United States.19 Follow-up is more limited 
for these patients than for the patient in our study, 
but initial reports in seven patients have not in-
cluded any new safety findings.19 Gene-transfer 
efficiency was lower than reported here, although 
therapeutic gene expression remained correlated 
with vector copy number values.
Outcomes in this patient provide further sup-
portive evidence to our previously reported results 
of patients who underwent a similar ex vivo gene 
therapy procedure for β-thalassemia with the 
same BB305 vector22,23 or the previous HPV569 
vector.23,24 In addition to the patient with sickle 
cell disease described here, under this same clini-
cal protocol, 4 patients with transfusion-dependent 
β-thalassemia have received LentiGlobin BB305. 
These participants had no clinically significant 
complications and no longer require regular trans-
fusions.22 These findings are consistent with early 
results reported with 18 other patients with thal-
assemia who received LentiGlobin BB305 in clini-
cal study HGB-204.23 Longer follow-up is required 
to confirm the durability of the efficacy and 
safety profile observed, and data from additional 
evaluations of gene therapy in a larger cohort of 
patients to confirm the promise of gene therapy 
for sickle cell disease are lacking.
Supported by Bluebird Bio and by a grant to the Biotherapy 
Clinical Investigation Center from Assistance Publique–Hôpi-
taux de Paris and INSERM.
Disclosure forms provided by the authors are available with 
the full text of this article at NEJM.org.
We thank the staff at Necker Children’s Hospital for their 
important contributions to the care of the patient described in 
this article; our colleagues Zoubida Karim, Ph.D., of Université 
Paris Diderot, Laurent Kiger, Ph.D., and Marie Georgine Rakoto-
son, Ph.D., of Institut Mondor de Recherche Biomédicale, and 
Michel Bahuau of Centre Hospitalier Universitaire Henri Mon-
dor for their contributions to the study; Laurent Kiger for creat-
ing and analyzing the oxygen-binding curves; Marie Georgine 
Rakotoson for creating and analyzing the density curves; Michel 
Bahuau for assessing the patient’s enzyme levels; Frédéric Ga-
lacteros for providing data on normal enzyme levels in patients 
with sickle cell disease; Mohammed Asmal, M.D., Ph.D., David 
Davidson, M.D., Tara O’Meara, Lilian Yengi, Ph.D., and Philip 
Gregory, Ph.D., of Bluebird Bio for their contributions to the 
study design and execution and for their critical review of an 
earlier version of the manuscript; and Katherine Lewis, an em-
ployee of Bluebird Bio, for editorial support in preparation of an 
earlier version of the manuscript.
Appendix
The authors’ affiliations are as follows: the Departments of Biotherapy (J.-A.R., A.M., E.M., L.C., M.C.), Clinical Pharmacy (P. Bourget), 
Pediatric Neuroradiology (D.G.), General Pediatrics (M.M.), and Pediatric Immunology–Hematology Unit (B.N., S.B.), Necker Chil-
dren’s Hospital, Assistance Publique–Hôpitaux de Paris (AP-HP), Biotherapy Clinical Investigation Center, Groupe Hospitalier Univer-
sitaire Ouest, AP-HP, INSERM (J.-A.R., A.M., E.M., L.C., L.W., M.C.), Unité de Technologies Chimiques et Biologiques pour la Santé, 
Centre National de la Recherche Scientifique Unité Mixte de Recherche 8258, INSERM Unité 1022, Faculté de Pharmacie de Paris, 
Université Paris Descartes, Chimie ParisTech (S.H.-B.-A.), Immunology Laboratory, Groupe Hospitalier Universitaire Paris-Sud, Hôpital 
Kremlin-Bicêtre, AP-HP, Le Kremlin-Bicêtre (S.H.-B.-A.), the Institute of Emerging Diseases and Innovative Therapies, Imagine Insti-
tute, Université Paris Descartes, Sorbonne Paris Cité University (M.S., B.N., L.W., M.C.), Mère-Enfant Clinical Investigation Center, 
Groupe Hospitalier Necker Cochin (M.S.), Université Paris Diderot, Sorbonne Paris Cité University, INSERM Institut National de Trans-
fusion Sanguine, Unité Biologie Intégrée du Globule Rouge, Laboratoire d’Excellence GR-Ex (W.E.N.), and Laboratoires de Virologie, 
Hôpital Cochin (J.-F.M.), Paris, Atomic and Alternative Energy Commission, Université Paris-Sud, Fontenay-aux-Roses (E.P., Y.B., S.C., 
P.L.), Institut Mondor de Recherche Biomédicale, Equipe 2, Centre de Référence des Syndromes Drépanocytaires Majeurs, Centre Hos-
pitalier Universitaire Henri Mondor, AP-HP, Laboratoire d’Excellence GR-Ex, Créteil (P. Bartolucci), and Université Paris Diderot, Sor-
bonne Paris Cité University, INSERM Unité 1149, Hôpital Louis-Mourier, AP-HP, Laboratoire d’Excellence GR-Ex, Colombes (H.P., T.L.) 
— all in France; Bluebird Bio, Cambridge (R.W.R., O.N., G.V., L.S., S.S.), and Brigham and Women’s Hospital and Harvard Medical 
School, Boston (P.L.) — both in Massachusetts; and Ramathibodi Hospital, Mahidol University, Bangkok, Thailand (P.L.).
The New England Journal of Medicine 
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 Copyright © 2017 Massachusetts Medical Society. All rights reserved. 
n engl j med 376;9 nejm.org March 2, 2017 855
Brief Report
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The New England Journal of Medicine 
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 Copyright © 2017 Massachusetts Medical Society. All rights reserved.